Mass spectrometers generally embody two primary components, an ion source that transforms species to be analyzed into ions in vacuum, and a mass analyzer that "weighs" these ions by measuring the response of their trajectories to some combination of electric and magnetic fields. For many years after their introduction, the use of such mass spectrometers was limited to species that were sufficiently volatile to be vaporized so that their dispersed molecules could be ionized by gas phase encounters with electrons, photons, other ions or electronically excited atoms. Non-volatile species could not be turned into ions by such gas phase encounters and therefore could not be individually weighed by ion mass analyzers. Thus, a long standing challenge in chemical analysis has been to extend the powers of mass spectrometric analysis to species that were so large and/or fragile that they could not be vaporized without substantial, even catastrophic, decomposition. In response to this need a number of methods have been developed in the past decade or two that are remarkably capable of transforming intact molecules of complex and fragile non-volatile species from a condensed phase into ions in vacuum ready for mass analysis. Among the most powerful and popular of these "soft" techniques is so called "Electrospray (ES) Ionization." It has been described at length in several U.S. Pat. Nos. (Labowsky et. al., 4,531,056; Yamashita et. al., 4,542,293; Henion et. al., 4,861,988, and Smith et. al., 4,842,701 and 4,885,706) and recent review articles [Fenn et. al., Science 246, 64 (1989); Fenn et. al., Mass spectrometry reviews 6, 37 (1990); Smith et. al., Analytical Chemistry 2, 882 (1990)]. To put the present invention in an perspective it will be appropriate to provide the following brief description of the method.
ES ionization as now practiced generally consists in flowing sample liquid comprising analytical species in a volatile solvent, at a rate typically between 1 and 20 microliters per minute, through a small metal capillary tube into a chamber containing bath gas at or near atmospheric pressure. An electrostatic potential difference of several kilovolts between the needle and the walls of the surrounding chamber produces an electric field at the needle tip that disperses the emerging liquid into a fine spray of charged droplets. Driven by the field toward the end wall of the chamber the droplets shrink by evaporation of solvent and, by a mechanism not yet clearly understood, produce free ions of the solute species in the ambient bath gas. Some of these ions become entrained in a small flow of bath gas through an aperture in the end wall leading into a vacuum system containing a mass analyzer. A most attractive feature of the ES technique is its "softness." Even the most fragile and complex species undergo little or no decomposition. Another attractive feature is that when the solute species comprise large polar molecules, for example biopolymers such as proteins, the ions produced by ES are multiply charged, roughly in proportion to their molecular weight. The result is that their mass/charge (m/z) values are usually less than 2500 or thereabouts so that they can be weighed by relatively inexpensive analyzers. Thus, ES ions of proteins with molecular weights of 130,000 have been accommodated by simple quadruple mass filters because they carry 100 or more charges. Indeed, from intact poly (ethylene oxide) oligomers with molecular weights up to 5,000,000 ES has produced ions with 4,000 or more charges!
In spite of these successes the ES technique is not without problems. Since the work of Vonnegut and Neubauer (Journal of Colloid Science 7, 616 (1952) it has been realized that stable sprays are difficult and sometimes impossible to obtain at atmospheric pressure with solutions having high electrical conductivities, especially when the solvent is mostly water. Moreover, the quality of the spray deteriorates when flow rates through a single needle exceed about 15 to 20 microliters/minute, depending upon the liquid properties. This loss of quality is reflected in poor spray stability and droplets that are too large and polydisperse in size. Another problem stems from the fact that with lower conductivity solutions the total spray current, i.e. flux of charge, is almost independent of the liquid flow rate. Consequently, increasing the flow rate decreases the charge/mass ratio in the droplets formed and, therefore, analytical sensitivity. These constraints complicate the use of ES sources with upstream sample processing such as liquid chromatography (LC) and capillary zone electrophoresis (CZE). Such separation techniques, and other sample treatments that may be desirable, have their own requirements for liquid composition so that compatibility with ES ionization can become difficult to achieve unless the ES step can be made less restrictive in its requirements.
Methods have been developed and applied that can overcome some of these difficulties. The teachings of the Henion patent mentioned above resurrect a technique tried years ago by Malcolm Dole's group [L.L. Mach et. al., Journal of Chemical Physics, 52, 4977 (1970)] and a technique used by the Fenn group from Yale University as shown in U.S. Pat. No. 4,531,056 listed above. Dole's group passed an aspirating gas in an annular flow around the liquid emerging from the injection needle in the hope that ES performance might be improved. They found instead that ion currents decreased so they abandoned further use of the technique. The Fenn group passed an annular gas flow around the liquid emerging from the injection needle to help stabilize the Electrospray particularly in the negative ion producing mode. Nearly 20 years after Dole's work, Henion's group discovered that this "pneumatic assist" allowed them to obtain adequately stable sprays at liquid flow rates as high as 100 or so microliters/minute, higher than unassisted ES can be run with optimal performance. Although these high liquid flow rates exact a penalty in terms of decreased sensitivity, this cost is sometimes sufficiently offset by the convenience of being able to work with sample solutions at flow rates encountered in liquid chromatographic separations. It also turned out that this pneumatic assist could provide acceptable spray stability with liquids that present difficulties in unassisted ES because properties such as electrical conductivity or surface tension are outside the range of optimal performance.
Another strategy that can be used for overcoming composition problems in the liquid is the "sheath flow" technique for use with Capillary Zone Electrophoresis interfacing with Electrospray Ionization taught in the R. D. Smith patent mentioned above. It introduces a second liquid having appropriate properties in annular or sheath flow around a core flow comprising the sample liquid emerging from a dielectric tube. Rapid mixing of the sheath and core flows at the needle tip produces a liquid whose composition is such that it can give rise to a stable ES spray of acceptable quality without the need for any pneumatic assist. This sheath flow technique was extended to running higher conductivity solutions with high aqueous content used typically in liquid chromatography solutions by Whitehouse et. al. (J., proceedings of the 38th Conference on Mass Spectrometry and Allied Topics, P427, (1990)). However, the technique does not significantly extend the range of liquid flow rate.
Even though these approaches have substantially expanded the range of composition and flow rates in liquids, each has compromises in convenience or sensitivity or range of operation. The present invention provides another approach to the sprayability problem that seems to overcome many if not most of the difficulties encountered the techniques of the prior art. Our experimental studies of electrospray dispersion revealed that one of the possible reasons for the difficulties in spraying liquids with high conductivity was that such liquids show an enhanced stability against break up into droplets. Microscopic examination of the tip region of a small tube at a potential of several kilovolts relative to nearby surfaces shows that the meniscus of the emerging liquid assumes a shape approximately that predicted by G. I. Taylor (Proceedings of the Royal Society A313, 453 (1969)) and since known as a "Taylor Cone." Emerging from the tip of this cone is a small filament or jet of liquid that breaks up into tiny droplets within a few jet diameters when the liquid is well-behaved in the sense that it forms a stable spray. When the liquid is highly conductive it does not break up as rapidly. Indeed with the right combinations of voltage and flow rate we could make the jet persist for a distance of over ten centimeters. One possible explanation for this somewhat surprising behavior emerges from a consideration of what happens when the surface of a jet is perturbed by undulations. Rayleigh showed long ago (Proceedings of the Roy. Soc., 29, P.71, (1879)) that if the amplitude of such "varicose" waves exceeds a critical value determined by the properties of the liquid, the waves are amplified by surface tension forces until the jet breaks up into droplets. Indeed such surface wave growth is the generally accepted explanation for the widely practiced nebulization of liquids by ejecting them from an orifice or nozzle as a high velocity stream or jet. When the jet is formed from a highly conductive liquid whose surface is charged, it is to be expected that the effective mobility of surface charge will also be high so that the surface potential, due to the surface charge, should at all times have the same value everywhere on the surface. On the other hand, the local electric field associated with such surface potential increases wherever the radius of curvature of the jet surface may decrease. This local increase in electric field decreases the effective value of the local surface tension, thus, damping the growth of the surface waves and inhibiting the break up of the jet into droplets. This picture accounts for the difficulties encountered in the ES nebulization of highly conductive liquids by purely electrostatic forces. It also suggests that these difficulties might be overcome by somehow providing surface perturbations sufficiently strong to overcome the stabilizing effect of the surface tension. We believe that the role of the gas flow in pneumatically assisted ES is to provide just such perturbations. The present invention is based on our discovery that mechanical vibrations provide a particularly effective and convenient source of the needed perturbations. Aqueous solutions with high conductivity can easily be sprayed with this invention when used in conjunction with Electrospray ionization. Ion production performance and subsequent analysis using mass spectrometry is not compromised with solution liquid flow rates even over 400 ul/min.